SUMMARY ABSTRACT The persistence of dormant, drug-resistant cancer cells after primary surgery and chemotherapy is a major factor contributing to poor outcomes for patients with ovarian cancer. The goal of this proposal is to understand mechanisms that regulate and sustain tumor dormancy in human ovarian cancer and to identify drug targets that regulate survival of dormant, autophagic cancer cells. Our group has discovered ARHI (DIRAS3), a master switch that induces both autophagy and tumor dormancy. ARHI is an imprinted tumor suppressor gene that is downregulated by multiple genetic and epigenetic mechanisms in the majority of ovarian cancers.1-5 Re- expression of ARHI at physiological levels blocks cancer cell proliferation, inhibits motility, induces autophagy by multiple mechanisms, and establishes tumor dormancy. When the ARHI protein is re-expressed in human ovarian cancer xenografts, cell proliferation and angiogenesis are arrested indefinitely, and autophagy is induced. If ARHI expression is downregulated, human ovarian cancer xenografts become vascularized and grow promptly to kill their hosts.1 Treatment of dormant xenografts with chloroquine, a functional inhibitor of autophagy, dramatically delays tumor outgrowth after ARHI is downregulated, suggesting that autophagy is required to sustain dormant cancer cells in a nutrient-poor environment.6 These observations have provided the first inducible model for tumor dormancy in human ovarian cancer. The clinical relevance of the xenograft model is supported by our studies of the small, poorly vascularized nodules of dormant metastatic ovarian cancer cells found on the peritoneal surface at ?second-look? operations following primary surgery and chemotherapy. Ovarian cancer cells in histologically positive second-look specimens contain autophagosomes with punctate LC3 and ARHI in 81-84% of cases, compared to only 21-23% of primary ovarian cancers from the same patients.2 Whether ARHI-positive autophagic cancer cells found at second-look are selected by chemotherapy from pre- exiting subpopulations in the primary cancers, or whether ARHI-negative cancer cells adapt to nutrient-poor conditions by reversing epigenetic changes, re-expressing ARHI and undergoing autophagy, is not known and is a major objective of this proposal. Uniform treatment of ovarian cancer patients within the Department of Gynecologic Oncology and Reproductive Medicine at MD Anderson will permit us to answer this question by analyzing clonal architecture in untreated primary and positive second-look specimens using whole-exome sequencing. Paired specimens from the primary cancer and positive second-look will also permit us to explore mechanisms that arrest growth, prevent angiogenesis, and permit survival of dormant residual disease.